Abstract
Rac1b is a tumor-specific splice variant of the Rac1 GTPase that displays limited functional similarities to Rac1. We have shown previously a novel cross-talk between Rac1 and beta-catenin, which induces canonical Wnt pathway activation in colorectal cancer cells. This prompted us to investigate if Rac1b, frequently overexpressed in colon tumors, contributes to Wnt pathway dysregulation. We show that Rac1b overexpression stimulates Tcf-mediated gene transcription, whereas depletion of Rac1b results in decreased expression of the Wnt target gene cyclin D1. Reconstitution experiments revealed an important difference between Rac1 and Rac1b such that Rac1b was capable of functionally interacting with Dishevelled-3 (Dvl-3) but not beta-catenin to mediate synergistic induction of Wnt target genes. In agreement, Dvl-3 but not beta-catenin caused increased activation of Rac1b levels, which may explain the functional cooperativity displayed in transcription assays. Furthermore, we show that Rac1b negatively regulates E-cadherin expression and results in decreased adhesion of colorectal cancer cells. RNA interference-mediated suppression of Rac1b resulted in reduced expression of Slug, a specific transcriptional repressor of E-cadherin, and a concomitant increase in E-cadherin transcript levels was observed. Intriguingly, mutation of the polybasic region of Rac1b resulted in complete loss of Rac1b stimulatory effects on transcription and suppressive effects on adhesion, indicating the importance of nuclear and membrane localization of Rac1b. Our results suggest that Rac1b overexpression may facilitate tumor progression by enhancing Dvl-3-mediated Wnt pathway signaling and induction of Wnt target genes specifically involved in decreasing the adhesive properties of colorectal cancer cells.
Highlights
Rac1 is one of the most extensively studied members of the Rho family of small GTP binding proteins
We examined the effects of Rac1b knockdown on the regulation of an endogenous Wnt target gene www.aacrjournals.org cyclin D1 and observed f40% reduction (Fig. 1D)
Because its discovery as a splice variant overexpressed in colon and breast tumors, there has been considerable interest in characterizing the functions of Rac1b to understand how it may contribute to tumorigenesis in these tissues [16, 17]
Summary
Rac is one of the most extensively studied members of the Rho family of small GTP binding proteins. It regulates diverse cellular processes, which include actin cytoskeleton organization, membrane trafficking, proliferation, and gene expression [1,2,3,4]. An additional level of regulation exists for Rho GTPases, whereby Rho-GDP dissociation inhibitors (Rho-GDI) sequester GDP-bound forms in the cytoplasm, preventing their activation. Exhaustive characterization of this GTPase has revealed the pivotal role it plays in the genesis of many cancers. Deciphering the functional consequences of its expression and contribution to tumorigenesis has been challenging because Rac1b shows limited similarities to known Rac functions
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